Studies on the C3 acceptor molecule were extended from earlier work with Epimastigotes (Epi) of T. cruzi to culture derived metacyclic trypomastigotes (CMT) of T. cruzi. C3 binding studies showed that nearly five-fold less C3 bound to CMT than to Epi during incubation in serum. GP72 on CMT, which is the predominant surface-iodinatable constituent on this life cycle stage, is an inefficient acceptor for C3 deposition. This is in contrast to Epi for which GP72 is the preferential C3 acceptor during incubation in serum. We next examined the molecular basis for the difference in alternative pathway activation on Epi and CMT. First, the form of C3 fragments on Epi and CMT during serum incubation was examined. Approximately 3/4 of C3 on Epi following a 60 minute incubation was present as C3b, and the remainder was present as iC3b. In contrast, 85%-90% of C3 on CMT was present as iC3b. Purified factors B,D,P and C3 were used to deposit C3b on Epi and CMT and saturation binding studies of I125 B and I125 H were done on Epi and CMT bearing C3b. These studies showed that H binding curves were biphasic and nearly identical on both Epi and CMT, with high and low affinity populations. In contrast, although B binding was monophasic and of high affinity on Epi, B binding was biphasic and of markedly decreased affinity and extent on CMT. We have concluded from these studies that control of alternative pathway activation in Epi and CMT of T. cruzi is exerted at the level of B binding, and presumably reflects either the known difference in the C3 acceptor on Epi and CMT or the presence of regulatory molecules that differ between these two life cycle stages. Preliminary experiments are underway to investigate the interaction of C3 with promastigotes (P) of L. donovani. Results show that C3 deposits on P via antibody-independent alternative pathway activation, that C3 binds covalently to a high molecular weight parasite constituent, and the majority of C3 is present as C3b.